Targeting Toll-Like Receptor 4 To Modulate Neuroinflammation in Central Nervous System Disorders

Expert Opinion on Therapeutic Targets
ISSN: 1472-8222 (Print) 1744-7631 (Online) Journal homepage: https://www.tandfonline.com/loi/iett20
Targeting toll-like receptor 4 to modulate

neuroinflammation in central nervous system
disorders
Gunnar R Leitner, Tyler J Wenzel, Nick Marshall, Ellen J Gates & Andis
Klegeris
To cite this article: Gunnar R Leitner, Tyler J Wenzel, Nick Marshall, Ellen J Gates & Andis
Klegeris (2019): Targeting toll-like receptor 4 to modulate neuroinflammation in central nervous
system disorders, Expert Opinion on Therapeutic Targets, DOI: 10.1080/14728222.2019.1676416
To link to this article: https://doi.org/10.1080/14728222.2019.1676416
Accepted author version posted online: 03

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Publisher: Taylor & Francis & Informa UK Limited, trading as Taylor & Francis Group
Journal: Expert Opinion on Therapeutic Targets
DOI: 10.1080/14728222.2019.1676416
Targeting toll-like receptor 4 to modulate neuroinflammation in central
nervous system disorders
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Gunnar R Leitner 1, Tyler J Wenzel 1, Nick Marshall 1, Ellen J Gates 1 and Andis Klegeris 1*
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Department of Biology, University of British Columbia Okanagan Campus, Kelowna, British Columbia,
V1V 1V7, Canada
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Corresponding author:
Andis Klegeris, Department of Biology, University of British Columbia Okanagan Campus, Kelowna,
British Columbia, V1V 1V7, Canada,
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e-mail: andis.klegeris@ubc.ca
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Information Classification: General

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Abstract
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Introduction: Adverse immune activation contributes to many central nervous system (CNS)
disorders. All main CNS cell types express toll-like receptor 4 (TLR 4). This receptor is critical
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for a myriad of immune functions such as cytokine secretion and phagocytic activity of
microglia; however, imbalances in TLR 4 activation can contribute to the progression of
neurodegenerative diseases.
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Areas covered: We considered available evidence implicating TLR 4 activation in the following
CNS pathologies: Alzheimer’s disease, Parkinson’s disease, ischemic stroke, traumatic brain
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injury, multiple sclerosis, multiple systems atrophy and Huntington’s disease. We reviewed
studies reporting effects of TLR 4-specific antagonists and agonists in models of peripheral and
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CNS diseases from the perspective of possible future use of TLR 4 ligands in CNS disorders.
Expert opinion: TLR 4 specific antagonists could suppress neuroinflammation by reducing

overproduction of inflammatory mediators; however, they may interfere with protein clearance
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mechanisms and myelination. Agonists that specifically activate myeloid differentiation primary-
response protein 88 (MyD88)-independent pathway of TLR 4 signaling could facilitate
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beneficial glial phagocytic activity with limited activity as inducers of pro-inflammatory

mediators. Deciphering the disease stage-specific involvement of TLR 4 in CNS pathologies is
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crucial for the future clinical development of TLR 4 agonists and antagonists.
Key Words: Alzheimer’s disease, astrocytes, cytokines, microglia, neurodegeneration,

neuroinflammation, Parkinson’s disease, MyD88, toll-like receptors

Article Highlights
• Toll-like receptor (TLR) 4 antagonists could have beneficial effects in those central
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nervous system (CNS) diseases that are accompanied by neuroinflammation
• TLR4 antagonists could inhibit overproduction of inflammatory mediators and cytotoxins
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by glia.
• TLR 4 antagonists could have adverse CNS effects by inhibiting phagocytosis by glia,
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reducing protein clearance and interfering with myelination.
• Selective TLR 4 agonists could be beneficial by upregulating the phagocytic activity of
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microglia, leading to enhanced clearance of damaged tissue and abnormal protein
aggregates associated with several different CNS diseases.
• Significant adverse effects can be caused by TLR 4 agonists inducing secretion of
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inflammatory mediators and cytotoxins.
• Agonists that selectively activate the TLR 4 downstream myeloid differentiation primary-
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response protein 88 (MyD88)-independent pathway should be developed and further

studied since such agents could increase phagocytic activity of glia without significantly
upregulating glial cytokines and cytotoxins.
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1. Introduction
Toll-like receptors (TLRs) are a family of pattern recognition receptors (PRRs) that
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contribute to the activation of the innate immune system. The TLR family in humans includes
ten different receptors, all of which are transmembrane proteins [1]. TLRs recognize pathogen-
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associated molecular patterns (PAMPs), such as lipopolysaccharide (LPS), and damage-
associated molecular patterns (DAMPs), such as high-mobility group box 1 (HMGB1) [2]. TLR
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4 is different from all other TLRs since (1) it needs an accessory protein, myeloid differentiation
factor 2 (MD-2), to activate, and (2) in its activated state, TLR 4 engages two separate
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downstream signaling mechanisms: myeloid differentiation primary-response protein 88
(MyD88)-dependent and MyD88-independent pathways (Figure 1). TLR 4 activation pathways
have been extensively reviewed before [3–5]. In brief, TLR 4 agonists activate the receptor by
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interacting with MD-2 to form a complex with TLR 4. For example, the lipid A component of
LPS from Escherichia coli O111:B4 binds to MD-2 and activates TLR4 on human immune cells,
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such as monocytes and macrophages. Cluster of differentiation (CD) 14 is required for this
activation [6]. CD14-independent assembly of the TLR 4/MD-2 and agonist complex is also
possible; for example, UT12 antibody can activate TLR4 in murine macrophages that have CD14
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knocked out [7–9]. In the MyD88-dependent pathway, the adaptor proteins include
toll/interleukin-1 receptor (TIR) homology domain-containing adaptor protein (TIRAP),
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interleukin (IL)-1 receptor associated kinases (IRAKs) and tumour necrosis factor (TNF)
receptor associated factor (TRAF) 6. The engagement of adaptor molecules in the MyD88-
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dependent pathway initiates signal transduction that leads to the activation of mitogen-activated
protein kinases (MAPKs) and nuclear factor kappa-light-chain-enhancer of activated B cells
(NF-κB), which initiates the production of several different cytokines and chemokines. In the
MyD88-independent pathway, the recruited adaptor proteins include TIR domain-containing
adaptor protein inducing interferon (IFN)-β (TRIF), TRIF-related adaptor molecule (TRAM),
and TRAF3. The engagement of adaptor molecules in the MyD88-independent pathway initiates
signal transduction pathways that lead to the activation of interferon regulatory factor 3 (IRF3),

promoting the production of IFN-β and endocytosis of the TLR 4/MD-2 and agonist complex
(Figure 1). Both MyD88-dependent and MyD88-independent pathways have been shown to
regulate phagocytosis in several different types of immune cells [10,11]; however, neither of
these two pathways are absolutely required for phagocytosis since TRIF and MyD88 double
knockout (KO) dendritic cells exposed to LPS retain phagocytic activity [12]. Furthermore, it is
important to note that these pathways regulate apoptosis. For example, MyD88 has been shown
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to regulate apoptosis of monocytic cells [13], while TLR 4-induced apoptosis of murine
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macrophages is mediated by MyD88-independent pathways [14].
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Activation of TLR 4 is often described as requiring the presence of CD14 [3,12].
Antibody-based agonists of TLR 4, such as UT12, have been shown to activate MyD88-
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dependent and MyD88-independent pathways, as well as increase phagocytic activity of primary
macrophages from CD14 KO primary murine macrophages [9]. It is important to note that the
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specific moiety structure of TLR 4 agonists required to selectively activate MyD88-dependent or
MyD88-independent pathways is unknown, but certain molecules, such as R-form LPS, can
specifically activate the MyD88-dependent pathway in CD14-deficient macrophages [15].
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Targeting TLR 4 with agonists or antagonists, or modulating its downstream signaling

pathways, may have a therapeutic potential in treating neurodegenerative diseases. In the brain,
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TLR 4 is expressed by neurons as well as the non-neuronal glial cells, which include microglia,
astrocytes, and oligodendrocytes. TLR 4 is expressed primarily by microglia, and to a lesser
extent by astrocytes, oligodendrocytes, and neurons [16,17]. Microglia are representatives of the
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mononuclear phagocyte system in the brain, and TLR 4 activation regulates some of their
functions, such as phagocytic activity [3–5]. TLR 4 agonists induce the secretion of
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inflammatory chemokines and cytokines, such as TNF-α, IL-1β, IL-6, IL-8, and MCP-1, by
microglia and astrocytes [3–5]. Since these pro-inflammatory molecules contribute to the
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pathogenesis of a range of neurological diseases, therapeutic agents targeting TLR 4 are

currently at various stages of preclinical and clinical development. This article discusses the
implications of modulating the activation of TLR 4 in the following central nervous system
(CNS) pathologies: traumatic brain injury (TBI), ischemia, Alzheimer's disease (AD),
Parkinson’s disease (PD), multiple sclerosis (MS), Huntington’s disease (HD), and multiple

system atrophy (MSA). Modulating the activation of TLR 4 can decrease inflammation and
enhance removal of the protein aggregates associated with these CNS pathologies.
In order to study the effectiveness of TLR 4 ligands, rodents are widely used to model
human peripheral and CNS disorders. It is important to note that the clinical translation of data
obtained in rodent models will face added challenges specific to TLR 4 signaling, in addition to
the well-documented differences between rodent and human innate and acquired immune
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responses [18]. Specifically, the ligand-recognition domain of TLR 4 is significantly different
between humans and mice [19]; therefore, agonists or ligands that bind to murine TLR 4 may not
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bind to human TLR 4. Furthermore, mice and human cells express TLR 4 differently [20]. For
example, in humans, both microglia and astrocytes express TLR 4; however, in mice, microglia,
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but not astrocytes, have been shown to express TLR 4. This could lead to different neuroimmune
responses to TLR 4 activation. An additional challenge facing clinical development of some of
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the TLR 4-specific agonists and antagonists is their bulky molecular structures, which could
prevent their crossing the blood-brain barrier (BBB).
Very few studies have assessed the CNS effects of TLR 4 agonists or antagonists
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directly; therefore, this review also describes the effects of TLR 4 agonists and antagonists in the
periphery, which could be relevant to modulation of the CNS physiological and pathological
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processes. First, peripheral immune responses, which can be induced by TLR 4 ligands, lead to
elevated serum levels of inflammatory cytokines. Since some of these mediators directly cross
either healthy or compromised BBB, they could interfere with the health of neurons and glia
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[21]; therefore, inhibiting peripheral immune responses by TLR 4 antagonists could have
beneficial CNS effects. Second, the ligand-binding and the intracellular signaling domains of
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TLR 4 are conserved across different cell types within a species [19]; therefore, antagonists or
agonists that bind to the TLR 4 on peripheral cells are likely to bind the TLR 4 of CNS cells and
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modulate their function if they are able to cross BBB.
We searched major databases including MEDLINE, PubMed Central, Science Citation

Index, and Google Scholar using keywords “toll-like receptor 4”, “antagonist”, “agonist”,
“ligand”, “cytokines”, “microglia”, “astrocytes”, “macrophages”, “peripheral nervous system”,
“central nervous system” and their variations between 7 May 2018 and 24 February 2019. We
identified additional relevant articles by researching the reference lists of the articles revealed by

the initial database searches. We identified over 400 papers, which were then studied to assess
their relevance to the main topic of this review. We focused on TLR 4-selective molecules that
could be used to modulate the secretion of inflammatory mediators by immune cells, modify the
phagocytic activity of immune cells, or reduce symptoms in animal models of CNS diseases.
Human clinical studies were not considered. The scope of this review was also limited to
therapies that are selective to TLR 4; therefore, molecules targeting other members of the TLR
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family, as well as drugs such as naloxone, which binds to both TLR 4 and opioid receptors, were
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not included. This led us to the subset of articles referenced in Tables 1, 2, 3, and 4.
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2. TLR 4 activation contributes to CNS pathologies
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TLR 4 can be activated by various endogenous DAMPs in addition to pathology-
associated proteins such as aggregates of amyloid-β peptides (Aβ) or α-synuclein [22]. All these
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structures bind TLR 4 and activate downstream signaling pathways in glia, inducing secretion of
reactive oxygen species (ROS) and cytokines such as IL-1β and TNF-α, which can lead to
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damage and death of neurons [23–26]. Neuronal death is accompanied by the release of DAMPs
into the extracellular space, which can then further activate TLR 4 [27]. On the other hand,
activation of microglial TLR 4 may improve clearance of neurotoxic proteins, such as Aβ and its
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aggregates, through enhanced phagocytic and autophagic activity [28]. Overall, however, chronic
TLR 4 activation is believed to be associated with glia-mediated neuronal death due to excessive
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secretion of cytotoxins [23,29]. Section 2 of this review outlines evidence implicating TLR 4
activation playing a pivotal role in the initiation, propagation, and progression of
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neurodegenerative diseases and other CNS pathologies.

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2.1. TLR 4 regulates cellular functions and pathophysiological processes of the CNS
Cells of the CNS, including microglia, astrocytes, and neurons, demonstrate pleiotropic
responses to TLR 4 stimulation. TLR 4-specific agonists cause glial cells to adopt altered
morphology and increase their proliferation rate as well as immunoreactivity for activation
markers [30]. In human microglia and astrocytes, TLR 4 activation causes the secretion of pro-
inflammatory cytokines, such as TNF-α, IL-6, and IL-1β [26,30,31]. TLR 4-mediated cellular

responses could accelerate pathology caused by neuroinflammation, astrogliosis, and
excitotoxicity, as well as impair the injury-repair mechanisms within the CNS. However, TLR 4
has also been demonstrated to enhance neuro-supportive functions conferred by astrocytes, such
as neurotransmitter recycling, as well as glial-mediated protein clearance pathways, such as
phagocytosis [32,33]. In neurons, TLR 4 activation increases action potential conductance
through the downstream cyclic adenosine monophosphate (cAMP)-dependent protein
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kinase/cyclic AMP response element binding protein (CREB) pathway; it also increases the
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proliferation rate of neuronal precursor cells [34,35]. Below we consider the TLR 4-mediated
effects on neuroinflammation, astrogliosis, excitotoxicity, and myelination, since modulating
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these processes by specific TLR 4 agonists or antagonists could have therapeutic benefit in
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several CNS disorders.
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2.1.1. TLR 4 activation contributes to neuroinflammation
Neuroinflammation is arguably the most destructive component of neurodegenerative

processes initiated in part through TLR 4 activation. Disease-specific accumulation of proteins
such as Aβ in AD, α-synuclein in PD, or DAMPs released from damaged and dying cells, such
as HMGB1, activate TLR 4 to initiate the neuroimmune response [26,36]. Neuroinflammation
can be considered pathogenic when the inflammatory activation of microglia and astrocytes
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becomes excessive or chronic, causing a sustained release of cytotoxins, such as TNF-α and
ROS, into the extracellular space of the CNS [37]. Excess extracellular cytotoxins lead to
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neuronal death. For example, TNF-α activates TNF-α receptors on neurons to initiate caspase-
dependent apoptosis and dysregulates glutamate signaling, which leads to excitotoxic death of
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neurons [38,39]. ROS and other cytotoxins secreted by microglia also can damage neurons and
contribute to neurodegeneration [40–42]. an
In TBI and ischemic stroke (IS), TLR 4-mediated responses induce and sustain
neuroinflammation, which is secondary to initial injuries. Following the initial insult of ischemia
or trauma, brain cells enter into a state of metabolic stress, as evidenced by perturbations in
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adenosine triphosphate (ATP), lactate, and ionic balances [43,44]. These stressors cause neuronal
damage, which enables the release of DAMPs leading to neuroinflammation [43]. For example,
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HMGB1, a TBI-associated DAMP, is released into the extracellular space by damaged cells
where it binds to and activates TLR 4 [45]. This relationship between trauma-induced DAMPs,
TLR 4, and inflammation was illustrated in rat models of TBI where trauma was followed by
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increases in IL-1 β and TNF-α protein expression, which was attenuated by silencing TLR 4
[46]. Further, a study investigating acute cerebral infarction in an animal model of IS found that
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TLR 4 mRNA and protein expression were upregulated proportionally to the severity of infarct
[47]. Together, these findings implicate TLR 4 as a receptor activated by trauma- and ischemia-
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related DAMPs responsible for initiation of neuroinflammatory reactions in these pathologies.

Thus, blocking TLR 4 activation might be beneficial in reducing sustained pathophysiological
changes that follow the initial damage of TBI or IS.
In AD, the abnormal accumulation of Aβ contributes to neuroinflammation by directly

activating TLR 4 on microglia and astrocytes, as well as by modulating neuronal expression of
TLR 4 [26,37,48]. Fibrillar Aβ increased the release of pro-inflammatory cytokines IL-1β, IL-6,

and TNF-α by microglia and astrocytes in a TLR 4-dependent manner [26,31]. TLR 4
overexpression was induced upon stimulation with Aβ oligomers in senescent rat neurons, which
effectively sensitized the cells to subsequent TLR 4 stimulation [49]. Taken together, these
findings suggest that Aβ can induce neuroinflammation and cause neurons to become more
susceptible to aberrant TLR 4 signaling. Thus, TLR 4 modulation may slow or suppress
neuroinflammation in AD and the associated neuronal death.
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In PD, α-synuclein induces TLR 4-dependent pro-inflammatory responses by glial cells,
which cause neuronal death [50]. Treatment of primary murine microglia and astrocytes with α-
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synuclein increased their secretion of pro-inflammatory cytokines and ROS compared to cells
with the TLR 4 gene knocked out [31]. These glial secretions can cause neuronal damage and
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death, which could accelerate the progression of PD symptoms. Therefore, inhibition of TLR 4
may be beneficial as treatment of neuroinflammation associated with PD and other
synucleinopathies.
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In MS, TLR 4 regulates the autoimmune responses that cause neuroinflammation and
neuronal death. Adaptive immune T helper cells that express IL-17 (Th17) and IL-1 (Th1) are
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implicated as having an early and potentially causative role in MS due to their ability to cross the
blood brain barrier (BBB). Once within the brain tissue, Th17 and Th1 cells recognize myelin
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proteins and become activated in a TLR 4-dependent manner, which leads to the secretion of the
pro-inflammatory cytokines IL-6 and IL-23 [22,51]. Microglia are stimulated by cytokines
secreted by activated Th17 and Th1 cells, which causes them to engage in neuroinflammatory
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reactions that damage and kill neurons [52]. Furthermore, microglia are transformed by Th17
cells to act as antigen presenting cells, which increases the targeted attack against myelin,
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leading to increased cytokine secretion by adaptive immune cells [53]. Since TLR 4 is required
for the initial recognition of myelin by T helper cells, suppressing TLR 4 activation may reduce
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the inflammatory responses of Th17 and Th1 cells [54]. For example, in experimental
autoimmune encephalomyelitis (EAE) models of MS, Th1 cells were shown to be primed
through TLR 4-dependent mechanisms. However, this study also found that stimulating TLR 4
reduced the cytokine secretion by Th17 cells exposed to myelin [55]. Similar cell type-specific
effects were described by Marta et al. [52], who illustrated that TLR 4 activation specifically
regulated cytokine release by dendritic, Th17, and Th1 cells, as well as induced neuroprotective

activity of microglia in EAE models [52]. Thus, modulating TLR 4 activation would be expected
to have complex effects on neuroinflammation associated with MS.
In HD, mutant huntingtin (mHTT) causes neuronal death leading to release of DAMPs,
which likely activate TLR 4, thus contributing to neuroinflammatory damage [56]. The possible
mechanisms by which mHTT causes neuronal death is reviewed by Estrada et al. [57].
Involvement of TLR 4 in HD was only recently highlighted in a study by Griffioen et al. [58],
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which demonstrated that in a murine model of HD, TLR 4 knockdown and KO mice had
increased lifespan [58]. This indicated that suppression of TLR 4 might be beneficial for
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reducing the damage caused by neuroinflammatory responses in HD. However, more research is
required to elucidate the mechanism by which TLR 4 suppression contributes to the lengthening
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of lifespan in this HD model, including the effects of TLR 4 activation on neuroinflammation in
HD.
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2.1.2. TLR 4 regulates astrocyte functions
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Recently, reactive astrocytes have garnered attention for their role in neuroinflammation
and for contributing to irreparable scarring of nervous tissues [59]. Stimulation of astrocyte TLR
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4 results in their increased proliferation and augmented secretion of cytokines and other
neuromodulating molecules such as neurotrophic factors [32,60]. For example, astrocytes in a
cerebral ischemia mouse model became reactive, proliferated, and migrated to the site of injury
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[61]. TBI causes astrocytes to lose neuron supporting functions, which can lead to
neurotransmitter dysregulation causing excitotoxic neuronal death [62] (see 2.1.3). In MS,
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astrocytes migrate towards damaged neuronal axons [63]. Accumulation of astrocytes in injured
tissues could have neuroprotective effects, but their sustained activation would lead to scarring of
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tissues and exacerbation of neuronal death, as well as damage to cerebral white matter [59];
therefore, blocking astrocyte TLR 4 activation may reduce their reactivity and reduce scarring of
surrounding tissues.

2.1.3. TLR 4 activation contributes to excitotoxicity
In PD, TBI, and IS, neuronal death can be induced by high concentrations of
neurotransmitters, such as glutamate, in the extracellular space, which dysregulates neuronal
glutamate/N-methyl-D-aspartic acid (NMDA) signaling through a mechanism known as
excitotoxicity [64]. Prolonged NMDA receptor stimulation induces Ca2+ influx, which increases
the excitability of neurons, leading to their damage and release of DAMPs. For example, in an ex
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vivo rat model, HMGB1 release preceded cell death in neurons stimulated with NMDA [65].
Since HMGB1 and other DAMPs activate TLR 4, this receptor could initiate neuroinflammation
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associated with excitotoxicity and subsequent death of neurons. This hypothesis is supported by
studies with N9 and EOC 20 microglial cell lines. Treatment of these cells with LPS caused the
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binding of TLR 4 to NMDA receptors [66]. This observation indicates that TLR 4 stimulation
can enhance excitotoxicity by physically binding TLR 4 to NMDA receptors and activating them
[67]. Lastly, excitotoxicity and death of neurons were attenuated by glycyrrhizin, which binds to
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and blocks the effects of HMGB1 [65]. Therefore, TLR 4 modulation may attenuate
glutamate/NMDA-induced neuronal death associated with PD, TBI, and IS.
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2.1.4. TLR 4 regulates oligodendrocytes and myelination

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Demyelination of cerebral white matter is a neurodegenerative process that begins early

in the pathogenesis of MS and accelerates with disease progression. The process of
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demyelination leads to impaired electrical signaling of neurons and their death [68]. TLR 4
modulation may slow this process by reducing the destruction and encouraging synthesis of
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myelin by oligodendrocytes, thereby improving neuronal health. As MS progresses, myelin is

damaged at a gradually increasing rate, and oligodendrocytes inevitably fail to synthesize enough
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myelin to support neurotransmission, which leads to neuronal death [69]. Further, as astrocyte-
and microglia-mediated neuroinflammation progresses in MS, the health of oligodendrocytes
themselves becomes compromised and they lose their supportive functions, including the ability
to produce myelin [70]. Experimental evidence indicates that processes of remyelination and
demyelination occurring throughout remission and relapse of MS are influenced by TLR 4
activation [71]. For example, an in vitro study showed that oligodendrocyte death was induced
upon activation of microglial TLR 4, and this cytotoxicity was inhibited by silencing TLR 4 [72].

Therefore, activation of this microglial receptor may play a direct role in oligodendrocyte death
and interfere with remyelination processes. However, in other models of nervous system injury,
activation of TLR 4 contributed to clearance and repair of myelin, and TLR 4 deficiency
impaired oligodendrocyte proliferation and differentiation [72,73]. Thus, TLR 4 agonists and
antagonists may offer a strategy for modulating oligodendrocyte function and myelination of
neuronal axons; however, more research is required to better understand the role different cell
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types play in this process. It is interesting to note that in AD, elevated levels of Aβ plaques and
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NFTs are associated with demyelination and oligodendrocyte impairment, but it is unknown if
TLR 4 is involved in this process [74].
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2.2. TLR 4 regulates clearance mechanisms, apoptosis, and formation of abnormal protein
deposits
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TLR 4-dependent mechanisms contribute to the CNS clearance processes, which include
phagocytosis and autophagy. During phagocytosis, extracellular material is engulfed to form
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phagosomes. This process terminates upon fusion of the phagosome with lysosomes that digest
the internalized molecules. Phagocytosis is comprehensively described by Botelho & Grinstein
[75]. Autophagy is the process by which cells degrade and digest organelles and intracellular
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proteins. Autophagy is further classified into three categories referred to as micro-autophagy,

macro-autophagy, and chaperone-mediated autophagy, which are comprehensively reviewed by
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Parzych & Klionsky [76]. Similar to phagocytosis, autophagic processes terminate with
lysosomal degradation of the target organelles and proteins. In this review, we consider the
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mounting evidence indicating that TLR 4 plays a pivotal role in phagocytosis performed by glial
cells, as well as the effects of TLR 4 activation on autophagy. Both these mechanisms may hold
relevance to designing therapeutic strategies for reducing pathologies associated with protein
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misfolding such as amyloidosis, synucleinopathies, and tauopathies.
2.2.1. TLR 4 mediates phagocytosis
Phagocytosis can be triggered by TLR 4 activation, and it enables the removal of large
structures such as cellular debris, proteinaceous plaques, and damaged or infected cells. In the

CNS, microglia act as the primary phagocytes with recent evidence indicating that astrocytes and
oligodendrocytes can also remove tissue debris by phagocytosis [77]. TLR 4 is one of the
principle receptors responsible for modulating phagocytosis [33]. For example, microglial TLR 4
deficiency leads to decreased phagocytosis, whereas chronic TLR 4 activation accelerates
clearance of myelin and other proteins, and can even cause premature engulfment of healthy
neurons [28,78]. Together, these data indicate that modulating microglial phagocytosis through
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TLR 4 activation may lessen protein accumulation and reduce secondary injury; however, this
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approach may also accelerate neuronal death by exacerbating neuroinflammation through
increased secretion of pro-inflammatory mediators and cytotoxins (see 2.1). Further research into
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TLR 4-mediated phagocytosis by astrocytes and oligodendrocytes is required for more complete
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understanding of the potential of TLR 4 modulators as enhancers of neurotoxic debris removal.
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2.2.2. TLR 4 regulates autophagy
Autophagy is a mechanism that facilitates removal of damaged organelles or abnormal

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intracellular aggregates of proteins associated with several CNS disorders including α-synuclein
in PD, or tau and Aβ in AD. TLR 4 ligands have been shown to either facilitate or inhibit
autophagy, which can have opposite effects on protein and organelle turnover. For example, low
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grade TLR 4 activation early in the disease process may stimulate autophagy of neurotoxic
proteins and damaged organelles within neurons, while chronic TLR 4 activation initiates
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premature organelle autophagy causing impairment of neuronal signaling and cell death [79]. In
the context of PD, α-synuclein dysregulates lysosomal function to reduce cellular autophagy and
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protein turnover. This in turn causes increased surface expression of TLR 4, a protein that is
primarily degraded by lysosomal activity, which likely affects a range of cellular functions [80].
Thus, activation of cellular autophagy by TLR 4 can be considered a feedback mechanism,
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which could influence other TLR 4-mediated responses.
2.2.3. TLR 4 regulates apoptosis of CNS cells
Apoptosis is a hallmark of many CNS diseases. It is well established that apoptosis of

neuronal cells contributes to neurodegeneration and the progression of cognitive symptoms in

TBI, IS, AD, and HD [43,58,81]. Further, microglial activation is controlled by TLR 4-
dependent apoptotic mechanisms, which may become dysregulated in CNS diseases [82].
Studies with microglia-like mononuclear phagocytes have demonstrated that PAMPs and various
endogenous DAMPs activate TLR 4 [3], and that this activation could induce neuronal apoptosis
directly or indirectly; TLR 4 activation induced TRIF-dependent apoptosis of murine
macrophages and also upregulated TNF-α secretion through MyD88-dependent mechanisms
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[13,14]. Elevated TNF-α, in turn, could induce apoptosis of both neurons and glia by interacting
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with their Fas/CD-95 death receptors [14]. In neurons, Aβ and peroxidised lipids were
demonstrated to cause TLR 4 activation, which induced c-Jun N-terminal kinase (JNK)- and
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caspase-3-mediated apoptosis [82]. The same study demonstrated that mutation of the TLR 4
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gene increased survival of neurons treated with Aβ; therefore, blocking TLR 4 activation may
help reduce apoptosis of neurons associated with neurodegenerative diseases, but inhibiting
apoptosis may adversely affect autoregulation of microglial activation preventing the removal of
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unwanted activated cells [83].
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2.2.4. TLR 4 enhances Aβ protein clearance
TLR 4 activation enhances the phagocytosis of extracellular Aβ aggregates, which may

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aid in the prevention and reduction of amyloidosis in AD, PD, TBI, and IS. In AD and PD,
extracellular aggregates and intracellular/cytoplasmic inclusions of Aβ are notable pathological
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features within brain tissue; these are also found in TBI and IS as the diseases progress [83,84].
In PD, Aβ deposition can exacerbate the damage caused by α-synuclein, which accelerates
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progression of cognitive symptoms [85,86]. Abnormal cleavage of amyloid precursor protein

(APP), altered ubiquitination, protein misfolding, and proteasomal dysfunction can contribute to
amyloidosis, which impedes cellular function, disrupts the BBB, and initiates cerebrovascular
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disease [87,88]. In mouse models of AD, TLR 4 activation has been demonstrated to reduce
amyloid plaque load by enhancing their clearance through phagocytosis by glial cells [28,29].
Therefore, activation of glial TLR 4 may enhance clearance of Aβ deposits not only in AD, but
also PD, TBI, and IS.

2.2.5. TLR 4 activation facilitates removal of α-synuclein
α-Synuclein plays a key role in neurodegenerative conditions known as

synucleinopathies, which include PD [89,90]. α-Synuclein facilitates its own accumulation by
causing prion-like misfolding of proximal proteins and interfering with the lysosomal and
proteasomal degradation of proteins [91,92]. Neuroinflammation impairs lysosomal protein
clearance pathways by decreasing cellular autophagy thereby further increasing intracellular α-
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synuclein accumulation and aggregation [46]. α-Synuclein is mainly intracellular, but when it
becomes externalized, α-synuclein can be toxic to neurons directly. It also activates glial TLR 4
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causing increased pro-inflammatory cytokine secretion, which may lead to indirect neurotoxicity
[93,94]. However, other studies have demonstrated that TLR 4 stimulation facilitates the removal
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of α-synuclein from the extracellular environment through, for example, enhanced phagocytosis
by microglia [50,78,89]. Therefore, agents modulating TLR 4 activity would be expected to have
complex effects on accumulation and clearance of α-synuclein.
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2.2.6. TLR 4 activation reduces tau accumulation
The activation of TLR 4 may reduce aggregation of hyperphosphorylated tau (p-tau)

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protein, a neurotoxin associated with AD, PD, and several other neurodegenerative conditions
[95]. Normally, tau protein participates in microtubule assembly and can be differentially
phosphorylated by various kinases in order to modulate microtubule stability [96]. While it is not
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clear why tau becomes hyperphosphorylated in AD, it has been established that
hyperphosphorylation makes tau more resistant to ubiquitination and proteasomal degradation,
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which leads to its aggregation and proteasome stress [88,91]. In PD, p-tau is found in high levels
in neurons and contributes to neurotoxicity by associating with Lewy bodies [97]. Interestingly,
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chronic TLR 4 stimulation has been demonstrated to reduce tauopathy in transgenic murine
models of AD [29]. Therefore, TLR 4 activity modulating agents could potentially be used to
attenuate tauopathy in neurodegenerative diseases.

3. Antagonists of MyD88-dependent and -independent signaling
TLR 4 activation can be prevented by blocking the transfer of an agonist, such as LPS,
from LBP to MD-2 (Figure 2). By inserting their long fatty acid tails into the binding cavity of
MD-2, antagonists, such as Eritoran, FP7, CRX-526, LPS from Rhodobacter sphaeroides (LPS-
RS), and LPS from cyanobacteria (CyP), prevent agonists, such as LPS from E. coli O111:B4,
from binding to the TLR 4/MD-2 complex [98,99]. Other antagonists, such as STM28 and TAK-
t
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242, bind directly to the extracellular and intracellular domain of TLR 4, respectively [100–102].
TLR 4 antagonists act by reducing dimerization of TLR 4/MD-2/agonist complexes, thus
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preventing TLR 4 activation, or by inhibiting activation of the downstream intracellular signaling
pathways by TLR 4/MD-2/agonist complexes. As previously mentioned, this review will only
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examine antagonists that are selective for TLR 4. Effects of TLR 4 antagonists in peripheral and
CNS diseases are summarized in Tables 1A and 1B, respectively.
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3.1. Effects in the periphery
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TLR 4 antagonists have been researched primarily in peripheral pathologies and have
shown promising therapeutic effects in models of diseases as diverse as septic shock, influenza
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virus infections, diabetic nephropathy, lung ischemia, vascular inflammatory disease, and
irritable bowel syndrome, among others [103–105]. Eritoran, STM28, and Vizantin have been
used in models of septic shock induced by Gram-negative bacteria. It has been hypothesized that
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LPS, which is a component of the outer membrane of Gram-negative bacteria, over-activates

TLR 4; therefore, certain TLR 4 antagonists could be effective in reducing the excessive immune
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response associated with sepsis and septic shock [100,106]. Importantly, Eritoran was shown to
be well tolerated by sepsis patients in a phase II clinical trial. Even though Eritoran did not
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improve clinical symptoms of septic shock compared to the placebo treatment, it showed a trend
towards reducing mortality associated with septic shock [98]. STM28 also effectively treated
endotoxin-induced sepsis in mice and reduced the secretion of inflammatory cytokines by human
THP-1 monocytes [100]. Clinical trials using STM28 have not been performed; therefore, safety
and efficacy studies with STM28 in humans are required. In vitro studies showed that Vizantin
reduced TNF-α and macrophage inflammatory protein (MIP)-1β secretion by LPS-treated THP-1

cells and attenuated the spread of Pseudomonas aeruginosa by inducing phagocytosis in
RAW264.7 macrophages [106–108].
Eritoran, FP7, and CRX-526 have been shown to reduce tissue damage caused by viral
infection in mouse models [103,109,110]. Treating influenza-infected mice with Eritoran or FP7
reduced TNF-α and IL-1β gene expression in lung tissue [104,109]. FP7 also reduced IL-8, IL-6,
IL-1β, and MIP-1α secretion in a study using C57/B6 mice treated with LPS [103]. FP7 reduces
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TLR 4 activation by binding to CD14 as well as MD-2 [111]. A study by Cluff et al. [112]
showed that CRX-526 did not affect resistance of BALB/c mice to H3N2 influenza infection;
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however, CRX-526 was protective in a mouse model of diabetic nephropathy [111] and reduced
symptoms in the dextran sodium sulfate (DSS) mouse model of inflammatory bowel disease
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(IBD) [105,110]. Epidemiological evidence implicates IBD as a risk factor for several
neurological diseases including AD and PD, and a recent study demonstrated that DSS model
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mice not only showed signs of peripheral inflammation (e.g., elevated serum IL-6), but also
developed neuroinflammation measured as increased cortical IL-6 and TNF-α levels, microglial
activation, and disrupted BBB [113]; therefore, controlling peripheral inflammatory conditions,
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such as IBD, by TLR 4 antagonists could have beneficial effects on neurological disorders driven
by neuroimmune mechanisms including glial activation.
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TAK-242, by blocking TLR 4 intracellular signaling, reduced symptoms of chronic

pancreatitis, such as pancreatic weight loss in Sprague-Dawley rats treated with trinitrobenzene
sulfonic acid [114]. In vitro studies showed that TAK-242 inhibited secretion of inflammatory
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mediators including TNF-α, IL-6 and nitric oxide (NO) by human U-937 monocytic cells and
mice peritoneal macrophages stimulated by LPS; however, the effect of this antagonist on
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cellular viability was not assessed [101,115]. Thus, preventing TLR 4 activation may have
beneficial effects in a range of peripheral diseases due to reduced levels of pro-inflammatory
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cytokines and NO.
LPS analogs derived from non-E. coli bacteria, such as LPS-RS and CyP, may also
reduce peripheral immune cell activation and cytokine secretion. For example, LPS-RS reduced
murine T cell proliferation and secretion of IFN-γ induced by LPS [116]. Further, CyP dose-
dependently reduced whole blood concentrations of IL-6, IL-8, IL-1β, and TNF-α in pigs treated

with ultra-pure LPS [117]. These studies indicate that LPS analogs could be used as
pharmacological interventions to reduce TLR 4-mediated peripheral inflammation.
3.2. Effects in the CNS
To date, only a few studies have considered the potential therapeutic benefits of
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preventing the activation of TLR 4 in CNS diseases (Table 1B). Elevated levels of pro-
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inflammatory cytokines including TNF-α, IL-1β, IL-8, and IL-6 have been reported in CNS
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diseases such as AD, PD, TBI and MS [118], which could be caused by TLR 4-dependent
mechanisms; for example, TLR 4 activation in TBI has been shown to be responsible for
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increased cytokine secretion, leading to chronic neuroinflammation and neurotoxicity [46].
Additionally, chronic TLR 4-dependent activation of microglia has been identified as a cellular
mechanism causing neuroinflammation and neurotoxicity in AD [4]. Therefore, targeting the
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TLR 4-induced microglial activation and secretion of pro-inflammatory cytokines could
potentially reduce neuroinflammation in these pathologies. As shown in Table 1B, several
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structurally different TLR 4 antagonists could be assessed for their ability to reduce
neuroinflammation associated with CNS diseases since they may lower pro-inflammatory
cytokine levels as well as inhibit adverse activation of glial cells; they include lipid A mimetics
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CRX-526, Eritoran, and FP7; a synthetic peptide known as STM28; and a derivative of trehalose
6,6′-dicorynomycolate called Vizantin.
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In addition, Gaikwad and Agrawal-Rajput [116] demonstrated that LPS-RS, a

lipopolysaccharide extracted from R. sphaeroides, reduced TNF-α and NO secretion by LPS-
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stimulated BV-2 mouse microglia cells. They also used co-cultures of LPS-stimulated BV-2
microglia and neuro2a mouse neurons to demonstrate that LPS-RS decreased BV-2-mediated
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neurotoxicity and inhibited microglial phagocytosis. Furthermore, immunofluorescence studies

showed that LPS-RS significantly reduced the nuclear translocation of p65 NF-κB in microglia
either directly or by inhibiting the upstream MAPK signaling [116]. These results indicated that
LPS-RS could be used to diminish neurotoxicity of immune-stimulated microglia. Similar
reduction in potentially neurotoxic secretions was observed with CyP, a form of LPS purified
from Oscillatoria planktothrix FP1, in neuron-microglia-astrocyte co-cultures derived from
C57BL/6J mice [119]. This study showed that CyP significantly lowered the release of TNF-α,

IL-6, and IL-1β from LPS-treated neuron-microglia-astrocyte co-cultures. In addition, CyP
reduced reactive astrogliosis and the expression of TNF-α in the spinal cord of Wobbler mice, a
model of amyotrophic lateral sclerosis (ALS). CyP also improved the negative symptoms of
motor neuron degeneration in these mice indicating that this TLR 4 antagonist could be
considered for further pre-clinical development as an ALS treatment option [119].
It is important to note that some TLR 4 antagonists, such as TAK-242 (a small molecule
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cyclohexane derivative) or FP7 (a glucosamine-based lipid A analogue), may also have harmful
effects in the CNS since they have been shown to reduce neuronal survival and proliferation
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[35,120]. While investigating the role of TLR 4 in the proliferation and differentiation of human
neural stem cells (hNSC), Grasselli et al. [35] observed that FP7 reduced the number of Ki67+
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proliferating cells and increased the number of Casp3+ apoptotic cells. The finding that FP7 is
detrimental to neurogenesis could prevent its use in neurodegenerative diseases, such as ALS and
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AD. Similarly, a study using TAK-242 to investigate the role of TLR 4 in brain recovery
following intracranial hemorrhage (ICH) in Sprague-Dawley rats found that this TLR 4
antagonist decreased restoration of neurological function and neurogenesis [120]. These adverse
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effects could limit the use of TAK-242 and FP7 in neurological diseases.
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4. Agonists and activators of MyD88-dependent and -independent signaling
TLR 4 activation mechanisms are summarized in Section 1 and Figure 1. Most agonists
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of this receptor bind to TLR 4/MD-2 causing dimerization of the complex leading to the
activation of downstream intracellular signaling pathways [121]. For example, lipid A mimetics
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E6020 and monophosphoryl lipid A (MPL) are vaccine adjuvants that selectively bind to TLR
4/MD-2 and initiate receptor activation independently of CD14 [9,122]. E6020 and MPL activate
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the MyD88-dependent signaling pathway causing increased production of pro-inflammatory

cytokines and chemokines in both the periphery and the CNS. MPL also activates the MyD88-
independent pathway [122–126].
Low molecular weight agonists such as 1Z105, 1Z88, and Euodenine also bind to MD-2
and activate both MyD88-independent and MyD88-dependent pathways without requiring CD14
[127–129]. 1Z105 and 1Z88 are non-lipid influenza vaccine adjuvants that were designed

specifically to activate TLR 4 [130]. UT12 is a monoclonal anti-TLR 4 antibody that binds to
TLR 4/MD-2 complex and induces MyD88-independent signaling and internalization of the
receptor complex [7]. Interestingly, Eritoran, a TLR 4-specific antagonist that binds MD-2, was
unable to block the dimerization and internalization of the receptor complex elicited by UT12.
This indicates that UT12 may bind an epitope on TLR 4/MD2 which is distinct from the LPS
binding site on MD-2 [8,9]. This section will discuss biological effects of agonists that
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selectively induce TLR 4 activation in models of peripheral and CNS diseases (Table 2A, 2B).
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4.1. Effects in the periphery
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The effects of TLR 4 agonists in peripheral tissues have been studied extensively with
most of the research to date conducted in the context of using TLR 4 agonists as vaccine
adjuvants [122,126]. While activation of TLR 4 commonly induces the secretion of cytotoxic
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molecules by immune cells, TLR 4 agonists have also been shown to enhance the clearance of
tissue debris and proteins. For example, MPL enhanced phagocytosis of Aβ by murine
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monocytes [124]. Therefore, activating specific downstream TLR 4 signaling mechanisms, such
as the MyD88-independent pathway, may preferentially enhance beneficial phagocytosis with
only moderately increased release of cytotoxins and pro-inflammatory mediators [124,131].
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Thus, treating murine CD8+ and CD4+ T cells with MPL preferentially induced the MyD88-
independent signaling pathway, which resulted in reduced cytotoxic secretions compared to cells
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stimulated with LPS [131]. E6020, another vaccine adjuvant, was also shown to stimulate
peripheral immune cells of rats in a TLR 4-specific manner, resulting in moderate cytokine
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secretion, low toxicity, and no adverse side effects [132]. Interestingly, MPL has been tested in
clinical trials as an anti-cancer vaccine adjuvant, and is being developed as an anti-Aβ vaccine
adjuvant [112,133]. Testing MPL in AD animal models has highlighted the TLR 4-mediated
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interplay between the peripheral and central immune responses. Administering MPL on its own
to AD model mice enhanced Aβ plaque phagocytosis, decreasing the number and the size of Aβ
deposits in their brains. This was attributed to direct activation of microglial TLR 4 by MPL. A
potentially beneficial effect of MPL, involving upregulation of peripheral immune responses,
was observed when this partial TLR 4 agonist was co-administered with Aβ, leading to increased
titre of anti-Aβ antibodies, which could facilitate Aβ clearance in the brain [133].

Using high-throughput screening, two small synthetic TLR 4-specific agonists, 1Z105
and 1Z88, were identified to activate TLR 4 in murine bone marrow-derived dendritic cells
(BMDCs) by inducing lower levels of inflammatory cytokines IL-1β and IL-12 p70 compared to
LPS-stimulated BMDCs [127,130]. These agonists appeared to be good candidates for further
immune tolerization studies since administration of 1Z105 or 1Z88 to mice prior to LPS
challenge decreased the sera concentrations of IL-6 25 and five fold, respectively, compared to
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mice not exposed to 1Z105 or 1Z88 [130]. Interestingly, arthritic mouse models treated with
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1Z105 or 1Z88 showed a significant reduction in arthritis-related symptoms, such as histological
signs of inflammation, bone erosion, and cartilage damage [127]. The unique ability of these new
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drugs to modulate peripheral immune cell activity specifically through TLR 4 activation
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indicates that they may hold promise for use as immune modulating drugs in the CNS.
The unique mechanism by which UT12, a monoclonal antibody, activates TLR 4 may
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explain the upregulated bacterial clearance without the associated increase in IL-6 or TNF-α
secretion by neutrophils in murine lung infection [134]. Further, peritoneal murine macrophages
pretreated with UT12 secreted significantly less TNF-α and IL-6 when subsequently challenged
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with LPS or a repeat exposure to UT12 [9]. These data indicate that UT12 treatment reduces
surface expression of TLR 4 by several different immune cell types, which could lead to
decreased inflammatory responses of these cells to subsequent exposure to other TLR 4 agonists
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including DAMPs. However, in vivo studies showed that UT12 treatment increased the
proliferation rate of peripheral B lymphocytes and macrophages, which suggests that this TLR 4
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agonist may increase peripheral immune system reactivity [8,135].
When compared to LPS-treated cells, peripheral blood mononuclear cells stimulated with
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Euodenine secreted less IL-12p40, IL-10, and TNF-α even though the level of TLR 4 stimulation
was similar as demonstrated by measuring the expression of a NF-κB reporter gene using a
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secreted alkaline phosphatase assay [128]. Importantly, Euodenine did not induce IL-5 secretion
by human peripheral blood mononuclear cells, which may allow use of this agonist in diseases
with already upregulated IL-5 levels, such as asthma [128,136]; however, more preclinical and
clinical studies with this compound are required.

4.2. Effects in the CNS
The activation of TLR 4 may be beneficial in certain neurodegenerative diseases. First,

low-grade activation of TLR 4 prior to a severe injury may reduce cytotoxic secretions by
immune cells, compared to cells exposed to immune insult without pre-exposure to TLR 4
agonists. For example, mice exposed to low doses of systemic LPS prior to an ischemic insult
showed less damage associated with inflammation, as well as reduced activation of microglia,
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and limited infiltration of neutrophils into the brain [137]. Further, when the initial exposure to a
TLR 4 agonist primarily activates the MyD88-dependent pathway, any subsequent re-activation
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of TLR 4 by LPS favours the MyD88-independent pathway [138]. This switch in signaling
pathways results in less cytotoxic secretions. For example, LPS-tolerized murine macrophages
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secreted more IFN-β, a molecule that reduces damage induced by ischemia in rabbits, compared
to cells exposed to LPS for the first time [138–140]. Therefore, exposing brain microglia to low
concentrations of TLR 4 agonists may reduce their subsequent secretion of neurotoxins. In
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addition, the activation of TLR 4 leads to increased phagocytic activity of microglia [124]. As
discussed in section 2.2.0, increasing the phagocytic activity of brain glial cells may aid in
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clearing the protein aggregates associated with AD, IS, PD, MS, MSA, or TBI.
To our knowledge, very limited research has considered use of TLR 4 agonists to slow
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down neurodegeneration (Table 2B). For example, the effects of 1Z105 and 1Z88 within the
CNS are unknown; however, they have been shown to reduce peripheral inflammatory
responses, which may be beneficial in CNS diseases such as AD [127]. It is important to note
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that TLR 4 agonists could also induce microglia-mediated CNS toxicity and neuronal damage;
however, certain agonists, such as E6020 and MPL when administered in vitro or in vivo did not
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elicit these adverse effects in rodent models of AD and MS. Multiple studies demonstrate that
these agonists could even aid in reducing symptoms associated with neurodegeneration
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[72,133,141]. Hypothetically, using specific agonists to cause a very selective activation of the
MyD88-independent TLR 4 signaling pathway may aid in the clearance of cytotoxic debris
within the CNS without causing excessive secretion of cytotoxins and inflammatory cytokines,
such as IL-6 or TNF-α [73].
An in vitro rat model of AD was used to show that treatment with MPL elicited
neuroprotective effects, which might operate by priming the neuroimmune system to sense and

remove Aβ oligomers before plaque deposition occurs in the extracellular space [141]. When
MPL was administered to mice pre-treated with Aβ oligomers, they exhibited improved
cognitive function, restored synaptic plasticity, and decreased Aβ deposits compared to control
mice not treated with MPL [133,141]. The neuroprotection provided by the MPL pre-treatment
was suggested to be mediated through the attenuation of downstream MyD88-dependent
signaling, which resulted in reduced TNF-α secretion and an increase in the production of anti-
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inflammatory mediators such as IRF3, IFN-β, and transforming growth factor (TGF)-β in the
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hippocampal region [142]. Therefore, MPL should be investigated further as a therapeutic option
for the early treatment and prevention of AD [133].
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E6020 and its CNS effects have been studied by Church et al. [73]; an in vivo MS model
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using an intraspinal lysolecithin injection to induce demyelination in rat spinal cords showed that
the co-injection of E6020 promoted axon sparing, clearance of myelin debris, and the infiltration
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of oligodendrocyte progenitor cells, which repair damaged myelin [73]. Further, advanced
preclinical studies using rats, rabbits, and cynomolgus monkeys showed little to no adverse
effects of E6020 [73,132]. While E6020 may hold promise as a therapeutic agent aiding
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remyelination in MS, more research is still required to determine its safety and efficacy in MS
patients. Although E6020 and MPL have shown promise as TLR 4 modulating therapies, they
have not gained any recent traction for further therapeutic research and development.
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5. Conclusions
The review of current literature reveals that, in addition to its already established role as a
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regulator of peripheral inflammation, TLR 4 controls several key physiological and

pathophysiological processes in the CNS. Presence and activation of TLR 4 and its downstream
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signaling pathways have been described in all major CNS cell types including glia and neurons.
TLR 4-dependent mechanisms are essential for neuroinflammatory responses, but also regulate
excitotoxicity, myelination, and clearance mechanisms including phagocytosis and autophagy.
All these mechanisms are common to several different CNS pathologies such as those considered
in this review: AD, PD, IS, TBI, MS, MSA, and HD. Therefore, drugs that selectively modulate
TLR 4 activity could be explored as therapeutic agents for these diseases. Only limited
preclinical studies using models of CNS disorders have been performed thus far using TLR 4-

specific ligands. Tables 1 and 2 illustrate that both TLR 4 agonists and antagonists show
beneficial or protective effects towards some CNS functions, while having potentially significant
adverse effects on others. Below we highlight the gaps in our knowledge revealed through
studies applying TLR 4 ligands in animal models of CNS diseases, and identify potential
avenues for future TLR 4-selective drug development.
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6. Expert opinion
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Regulating TLR 4 activation by using selective agonists and antagonists may provide a
new therapeutic approach for treatment of a wide range of diseases, including disorders of the
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CNS. Current therapies for neurodegenerative diseases, such as rivastigmine for AD and PD,
may ease some of their symptoms, but they do not slow disease progression and often possess
significant adverse effects [143]; therefore, preclinical development of new classes of drugs
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acting through previously unexplored cellular and molecular targets is essential.
The main beneficial activity of selective TLR 4 antagonists in neurodegenerative diseases

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could involve inhibition of inflammatory cytokine overproduction by glial cells, which
accompanies neuroinflammation associated with these disorders [144]. The key adverse activity
ed
of TLR 4 antagonists could be related to their inhibition of clearance of abnormal proteins and
cellular debris, as well as interference with myelination. Even though TLR 4 antagonists such as
STM28, Vizantin, and Eritoran have been shown to decrease cytokine production in the
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peripheral tissues, their ability to reduce CNS cytokine levels is unknown; thus, further studies
with these drugs using in vivo models of neurodegenerative diseases are needed. Other
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compounds such as LPS-RS, CYP, and Resatorvid (TAK-242) have been studied in several
models of CNS diseases, where they have shown promise by reducing neuroinflammation;
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therefore, performing more advanced studies of their pharmacokinetics and pharmacodynamics

would be the next logical step [102,106,109,116,119,120].
Agonists of TLR 4 may offer an alternative avenue for targeting some of the
pathophysiological mechanisms associated with certain neurodegenerative conditions. For
instance, observations that E6020, a vaccine adjuvant, promotes myelin regeneration in the
spinal cords of MS model rats provides evidence that this drug could be beneficial in MS [73].

MPL, another vaccine adjuvant, could potentially be developed as a therapeutic agent for AD, as
it has been shown to prevent the accumulation of Aβ in AD mice models [133]. Since TLR 4
activation would be expected to upregulate neuroinflammation and promote glial scarring [63],
further careful preclinical and clinical safety studies will be required before these drugs could be
recommended for clinical trials with patients suffering from neurodegenerative and other CNS
diseases. Design and optimization of agonists selectively targeting the TLR 4 MyD88-
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independent signaling pathway could be a possible future drug development strategy since such
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agonists may preferentially enhance beneficial phagocytosis while only moderately upregulating
pro-inflammatory mediators in both the periphery and the CNS [124,131].
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The effectiveness of treatments strategies targeting TLR 4 may depend on the stage of the
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disease. For example, the damage caused by neuroinflammation in TBI and IS may be prevented
by suppressing TLR 4 activation [46,145]; therefore, TLR 4 antagonists should be administered
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very soon after the initial damage in order to prevent the chronic injury that follows that acute
phase in these pathologies. Using TLR 4 agonists to prevent Aβ accumulation may be beneficial
only in the early stages of AD; therefore, early AD diagnosis will be essential for the TLR 4
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treatment to be effective. It should also be noted that suppressing neuroinflammation has been
proposed to be beneficial at any stage of PD and HD [90]. Deciphering the disease stage-specific
involvement of TLR 4 in different CNS pathologies is clearly an essential objective for future
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clinical development of TLR 4 agonists and antagonists as therapies for CNS diseases.
Funding
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This paper was not funded.

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Declaration of interest
The authors have no relevant affiliations or financial involvement with any organization or entity
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with a financial interest in or financial conflict with the subject matter or materials discussed in
the manuscript. This includes employment, consultancies, honoraria, stock ownership or options,
expert testimony, grants or patents received or pending, or royalties.
Reviewer disclosures
Peer reviewers on this manuscript have no relevant financial or other relationships to disclose

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Figures and Tables
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Figure 1. TLR 4 is activated after a complex of agonist with TLR 4/MD-2 is formed. Once
activated, TLR 4/MD-2 and agonist complexes dimerize and engage two separate downstream
signaling pathways: the MyD88-dependent and MyD88-independent pathways. The MyD88-
dependent pathway leads to the secretion of inflammatory molecules and increased phagocytic

activity, whereas the MyD88-independent pathway leads to the secretion of IFN-β and increased
phagocytic activity. IFN-β activates genes such as suppressor of cytokine signaling (SOCS)-1
and SOCS-3 in astrocytes [146]. Upon activation of the MyD88-independent pathway, the TLR
4/MD-2 and agonist complex becomes internalized by endocytosis. The exact mechanisms of
interaction between TRIF and the MyD88-dependent pathway are unknown, but it is established
that TRIF influences the MyD88-dependent secretory profile of immune cells. Both pathways
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have been shown to increase the phagocytic activity of immune cells, such as microglia.
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However, they may not increase phagocytic activity through the same mechanisms.
Abbreviations: CD14 = Cluster of differentiation 14; IFN = interferon; IL = interleukin; IRF3 =
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interferon regulatory factor 3; IRAK = IL-1 receptor associated kinases; LBP = lipid binding
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protein; LPS = lipopolysaccharide; MAPK = mitogen-activated protein kinase; MD-2 = myeloid
differentiation factor 2; MyD88 = myeloid differentiation primary-response protein 88; NF-κB =
nuclear factor kappa-light-chain-enhancer of activated B cells; TNF = tumour necrosis factor;
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TRAF6 = TNF receptor associated factor 6; TIRAP = toll/interleukin-1 receptor homology
domain-containing adaptor protein; TLR = toll-like receptor; TRIF = toll/interleukin-1 receptor
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domain-containing adaptor protein inducing IFN-β; TRAM = TRIF-related adaptor molecule.
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Figure 2. The activation of TLR 4 can be blocked by three mechanisms: (1) prevent binding of
the CD14-independent and CD14-dependent agonists to the TLR 4/MD-2 complex; (2) block
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dimerization of the TLR 4/MD-2/agonist complexes; (3) prevent activation of the downstream
intracellular signaling pathways by the TLR 4/MD-2/agonist complexes; Abbreviations: CD14 =
Cluster of differentiation 14; LBP = Lipid binding protein; LPS = Lipopolysaccharide; MD-2 =
pt
Myeloid differentiation factor-2; TLR 4 = Toll-like receptor 4.

ce
Table 1A. Biological effects of synthetic and natural TLR 4-selective agonists in the periphery
In Vitro Studies In Vivo Studies

Ac
Name(s) and Mechanism of Adverse Effects

structure Action
CRX-526 Binds to MD-2, N/A Studies using N/A

prevents agonist advanced diabetic
Lipid A
binding [105]. C57BL/6-Nos3t-
mimetic
m1Unc model mice.

derived from
aminoalkyl Results: CRX-526
glucosaminide reduced renal tubule
4-phosphate damage 1.2 fold in
advanced diabetic
model mice
t
compared to
ip
untreated mice [110].
cr
Studies using
us
BALB/cAnN mice
treated with DSS to
induce gut
an
inflammation.
M
Results: CRX-526
reduced gut
inflammatory disease
ed
activity in mice
exposed to DSS two
pt
fold compared to
control mice [105].
ce
Ac

Eritoran Binds to MD-2, N/A Studies using N/A
prevents agonist influenza virus
Lipid A binding [109]. infected C57BL/6
derivative mice.
Results: Eritoran
t
reduced influenza-
ip
induced lethality two-
cr
fold. Eritoran
reduced mRNA
us
expression of TNF-α
40 fold, and IL-1β 24
fold, in the lungs of
an
infected mice
compared to non-
M
infected mice [109].
FP7 Binds to MD-2, Studies using Studies using N/A

ed
prevents agonist monocytes from C57BL/6 mice.

Glucosamine- binding [104]. C57BL/6 mice.
based lipid A Results: FP7
pt
analog Results: FP7 protected mice from

inhibited LPS- influenza virus -
ce
induced secretion of induced lethality

IL-8, MIP-1β, and [104].
Ac
IL-6 compared to
monocytes not
treated with FP7
[104].

STM28 Binds to the TLR Studies using Studies using N/A
4 extracellular differentiated human C57BL/6 mice
Synthetic domain, prevents THP-1 monocytic treated with LPS as
peptide targets LPS-induced cells. an endotoxin shock
the activation [100]. model.
extracellular Results: STM28
t
domain of TLR reduced LPS-induced Results: STM28
ip
4 secretion of TNF-α decreased mortality
cr
six fold [100]. four fold compared to
controls exposed to
us
LPS only [100].
Studies using
an acetaminophen-
induced liver failure
in male C57BL/6
M
mice.
Results: STM28
ed
reduced TNF-α two

fold in the plasma,
and IL-1α two fold in
pt
the liver when

ce
compared to mice
treated with
acetaminophen only
Ac
[102].
TAK-242 Binds intracellular N/A Studies using N/A

(Resatorvid) TIR domain Sprague-Dawley rats
(Cys747 residue) treated with 2,4,6-
A small of TLR 4, trinitrobenzenesulfo-

molecule prevents activation nic acid to induce
cyclohexane of TLR 4 adaptor pancreatitis-like
derivative molecules [101]. symptoms.
Results: TAK-242
reduced symptoms of
t
chronic pancreatitis
ip
measured as a two-
cr
fold increase in
pancreas weight to
us
body weight ratio
an [114].
Vizantin Binds to MD-2, Studies using human N/A N/A

prevents agonist THP-1 monocytic
Derived from binding [106]. cells.
M
trehalose 6,6′-
dicorynomyco- Results: Vizantin
ed
late, a induced the secretion

component of of MIP-1β and IL-8.
the cell wall of THP-1 cells treated
pt
Mycobacterium with Vizantin prior to

tuberculosis LPS stimulation
ce
secreted five-fold less

IL-1β and TNF-α
Ac
compared to cells
stimulated in the
absence of Vizantin
[106].
LPS-RS Binds to MD-2, Studies using N/A N/A

prevents agonist splenocytes from

LPS purified binding [119]. c57BL/6 mice,
from exposed to LPS and
Rhodobacter treated with LPS-RS.
sphaeroides
Results: LPS-RS
ameliorated LPS-
t
induced T cell
ip
proliferation and
cr
IFN-γ secretion
[116].
us
CyP Binds to MD-2, Studies using porcine N/A N/A
prevents agonist whole blood exposed
LPS purified binding [119]. to LPS.
an
from
Oscillatoria Results: CyP
M
planktothrix treatment reduced
FP1 concentrations of
ed
TNF-α, IL-1β, and

IL-8 [117].
pt
Table 1B. Biological effects of synthetic and natural TLR 4-selective agonists in the CNS
ce
Name(s) and Mechanism of In Vitro Studies In Vivo Studies Adverse Effects

structure Action
Ac

FP7 Binds to MD-2, N/A N/A Studies using
prevents agonist hNSC and
Glucosamine- binding [104] primary human
based lipid A oligodendrocytes.
analog
Results: FP7
t
reduced the
ip
proliferation of
cr
hNSC cells and
oligodendrocytes
us
[35]
TAK-242 Binds intracellular N/A Studies using Studies using

(Resatorvid) TIR domain
an
hSOD1G93A Sprague-Dawley
(Cys747 residue) transgenic ALS rats modeling
A small of TLR 4, model mice. ICH.
M
molecule prevents activation
cyclohexane of TLR 4 adaptor Results: TAK-242 Results: TAK-
ed
derivative molecules [101]. reduced 242 reduced the

concentrations of recovery rate of
serum IL-1β two neurological
pt
fold, cerebrospinal functions after

fluid TNF-α 2.4 fold. ICH [120].
ce
Spinal astrogliosis
was reduced 2.5 fold
Ac
and microglial
activation 22.5 fold
in TAK-242-treated
ALS model mice
compared to mice not
treated with TAK-

242 [147].
LPS-RS Binds to MD-2, Studies using BV-2 N/A N/A

prevents agonist murine microglia and
LPS purified binding [119]. neuro2a murine
from neurons.
Rhodobacter
t
ip
sphaeroides Results: LPS-RS
decreased secretion
cr
of LPS-induced TNF-
α and NO secretion
us
2.5-fold compared to
microglia stimulated
an
in the absence of
LPS-RS. LPS-RS
increased neuron
M
survival four-fold in
LPS-stimulated
ed
microglia-neuron co-
culture, compared to
a microglia-neuron
pt
co-culture not treated

with LPS-RS. LPS-
ce
RS reduced
phagocytic activity of
Ac
LPS-stimulated BV-2
microglia two fold
compared to BV-2
cells not treated with
LPS-RS [116].

Studies using primary
neuron-glia co-
cultures from
C57BL/6J mice.
Results: LPS-RS
t
reduced the LPS-
ip
induced TNF-α and
cr
IL-6 secretion two
fold, and that of IL-
us
1β ten-fold compared
to neuron-glia co-
cultures not treated
an
with LPS-RS [119].
CyP Binds to MD-2, Studies using primary Studies using a N/A

M
prevents agonist neuron-microglia- wobbler mice model

LPS purified binding [119]. astrocyte co-cultures of ALS.
ed
from from C57BL/6J mice.

Oscillatoria Results: Treatment
planktothrix Results: CyP with CyP inhibited
pt
FP1. reduced TNF-α, IL- reactive astrogliosis,

1β, and IL-6 as well as reduced
ce
concentrations two- disease symptoms,

fold in LPS- such as poor grip
Ac
stimulated co- strength and paw

cultures, compared to abnormalities [119].
co-cultures not
treated with CyP.
CyP prevented death
of neurons induced

by exposure of co-
cultures to LPS
[119].
Abbreviations: ALS = amyotrophic lateral sclerosis; DSS = dextran sodium sulfate; CyP =
cyanobacterial LPS; hNSC = human neural stem cell; IL-1β = interleukin-1β; ICH =
intracranial hemorrhage; LPS = lipopolysaccharide; LPS-RS = lipopolysaccharide from
t
ip
Rhodobacter sphaeroides; MD-2 = myeloid differentiation factor 2; MIP-1β = macrophage
inflammatory protein-1β; N/A = not applicable since relevant data cannot be identified; NO =
cr
nitric oxide; TIR = Toll/interleukin-1 receptor; TNF-α = tumor necrosis factor alpha.
us
Table 2A. Biological effects of synthetic and natural TLR 4-selective agonists in the periphery
an
structure Action
M
E6020 and Binds TLR Studies using Studies using Study using rats
analogs 4/MD-2 human whole Balb/C female mice
Results: Two
independently of blood.
ed
Synthetic Results: IL-10 was weeks of

CD14 [132].
lipid A Results: TNF-α detectable in the subcutaneous
mimetic was detectable in serum of mice administration at
pt
the serum of subcutaneously 5 mg/kg/day

ER803022 (an injected with caused no
ce
E6020 analog) various E6020 observable

treated blood analogs [149]. adverse effects
Ac
samples [148]. other than

increased spleen
weight, white
Studies using the blood cell,
human glioma cell neutrophil, and
line U373 lymphocyte

Results: E6020 counts. These
analogs caused IL-6 effects returned to
secretion [148]. pre-
administration
levels within 24 h
Studies using [132].
t
human peripheral
ip
blood mononuclear
cells (PBMCs). Study using
cr
rabbits
Results: PBMCs
us
stimulated with Results: no
E6020 secreted 50- pyrogenicity or
fold more IL-1α and mutagenicity was
an
IL-1β, three-fold observed at a dose
more IL-2, 320-fold of 30 ng/kg [132].
M
more IL-6, 12-fold
more IL-10, 18-fold
more granulocyte- Study using
ed
macrophage colony- cynomolgus
stimulating factor, monkeys

pt
and 72-fold more Results: After

TNF-α compared to 0.01 mg/kg
ce
unstimulated cells subcutaneous

[132]. injection, no
Ac
cardiovascular or
respiratory
adverse effects
were observed; no
effect on body
temperature or

other clinical
signs.
0.1 and 1 mg/kg
subcutaneous
injection caused
swelling and
t
redness at the
ip
injection site.
1 mg/kg
cr
subcutaneous
us
injection
increased systolic
and diastolic
an
blood pressure,
heart rate, body
M
temperature, and
respiratory rate.
These effects
ed
diminished after
24 h [132].
pt
Monophosph- Binds MD-2 N/A Studies using N/A

oryl lipid A [131]. C57BL/6 mice.
ce
(MPL)
Results: Mice
Lipid A intravenously
Ac
derivative injected with MPL,

followed by an
intravenous injection
of amyloid-β peptide
(Aβ), had 12.5-fold
more Aβ internalized

by peripheral blood
monocytes,
compared to mice
injected with Aβ
alone [124].
Intraperitoneal
t
ip
injection of MPL
caused a 6000-fold
cr
increase in the serum
concentrations of
us
interferon-γ-induced
protein-10 and
an 23000-fold increase
in CXC motif
chemokine ligand
M
(CXCL)-1 compared
to control mice
[124].
ed
UT12 Binds TLR Studies using Studies using mice N/A

4/MD-2 complex murine peritoneal intranasally
pt
Antibody
to cause macrophages. coinfected with
developed
dimerization. Streptococcus
ce
against Results: UT12

murine Specific epitope pneumoniae and
induced TNF-α
tumour cells is unknown [7]. influenza virus.
secretion in
Ac
overexpressin macrophages from Results:

g mouse the wild-type and Intraperitoneal
TLR 4 injections of UT12
CD14 KO animals.
UT12 reduced LPS- reduced serum
induced secretion of concentrations of
TNF-α 2.5 fold TNF-α, IL-6,

compared to and keratinocyte-
peritoneal derived chemokine
macrophages two fold, as well as
stimulated in the MIP-2 1.5 fold,
absence of UT12 compared to control
[150]. mice [150].
t
ip
UT12 increased the
14-day survival of
cr
S. pneumoniae- and
influenza virus-
us
infected mice two
fold compared to
an infected mice not
treated with UT12
[150].
M
1Z105/1Z88 Bind TLR 4/MD- Study using Study using Study using
ed
2 independently primary murine C57BL/6 mice. HepG2 cells

Low
of CD14 [127]. bone marrow-
molecular Results: Mice Results: 1Z105
pt
derived dendritic
weight orally gavaged or and 1Z88 were
compounds cells (BMDCs).
intravenously not toxic to cells
ce
containing Results: 1Z105 and injected with 1Z105 at 10 µM [127].

pyrimido[5,4- 1Z88 blocked the or 1Z88 had 10-to-
Ac
b] indole secretion of IL-6 20-fold lower

scaffold
and TNF-α by LPS- serum Study using
structure
stimulated BMDCs concentrations of gastrocnemius
[127]. IL-6, TNF-α, and muscle harvested
interferon-γ- from BALB/c
induced protein-10 mice

Study using human compared to control Results: Minimal
myeloid dendritic mice [127]. inflammation at
cells. the injection site
Study using adult
of 1Z105 [130].
Results: 1Z105 arthritic K/BxN
induced secretion of mice.
IL-1β, IL-6, IL-8,
t
Results:
ip
IL-12 p70, and
Intraperitoneal
TNF-α by dendritic
injections of 1Z105
cr
cells. [130].
or 1Z88 reduced
paw swelling, paw
us
inflammation, bone
erosion, and
an
cartilage damage by
2.75 to five fold
compared to
M
arthritic mice not

treated with 1Z105
ed
or 1Z88 [127].
pt
Euodenine Binds TLR Study using human N/A Study using

4/MD-2 PBMCs. human PBMCs
ce
Isolated from
independently of
the leaves of Results: Euodenine Results:
Euodia CD14 [128].
induced secretion of Euodenine was
Ac
asteridula IL-8, IL-12P40, IL- toxic to cells at

(Rutaceae) 10, and TNF-α by concentrations
unstimulated above 30 µM
PBMCs, and blocked [128].
secretion of IL-5 by
LPS-stimulated

PBMCs. While
Euodenine induced
the secretion of IL-8
at levels comparable
to LPS treatment,
IL-12P40, IL-10,
t
and TNF-α levels
ip
were two- to three-
fold lower in
cr
Euodenine-
us
stimulated cells
[128].
an
M
ed
pt
ce
Ac

Table 2B. Biological effects of synthetic and natural TLR 4-selective agonists in the CNS

structure Action
E6020 and Binds TLR N/A Studies using adult Studies using adult
analogs 4/MD-2 female Sprague female Sprague
t
independently of Dawley rats. Dawley rats.
ip
Synthetic
CD14 [132].
lipid A Results: Intraspinal Results: Injection of
cr
mimetic injection of E6020 E6020 into rat spinal
into rats increased cords caused
us
macrophage oligodendrocyte loss
activation in white one day post-
matter four fold injection, followed
an
compared to mice by a significant
injected with increase in
M
phosphate-buffered oligodendrocyte
saline (PBS) alone number on day seven
[73]. post-injection.
ed
Occasional loss of
Intraspinal injection
neurons was
of E6020 increased
pt
observed in gray
the proliferation of
matter one day post-
oligodendrocyte
ce
injection, which was

progenitor cells eight
associated with
fold, as well as
robust macrophage
Ac
increased the
activation [73].
number of mature
oligodendrocytes six
fold compared to rats
injected with PBS
alone [73].

Intraspinal co-
injection of E6020
with lysolecithin (to
induce
demyelination)
increased the
t
removal of myelin
ip
debris by
macrophages ten-
cr
fold after 14 days,
us
compared to rats
injected with PBS
alone. Rats co-
an
injected with E6020
and lysolecithin had
M
3.5-fold lower
axonal loss
compared to mice
ed
injected with PBS

and lysolecithin [73].
pt
Monophospho Binds MD-2 Studies using BV-2 Studies using N/A

ce
ryl lipid A [131]. murine microglia APPSWE/PS1 mice

(MPL) cells. (AD model).
Ac
Lipid A Results: MPL Results:

derivative upregulated TNF-α Intraperitoneal
secretion 15 fold, as injections of MPL
well as the monocyte increased cognitive
chemoattractant test scores of AD
protein (MCP)-1 model mice 1.8 fold,

mRNA levels two as well as reduced
fold compared to the size and number
BV-2 microglia of Aβ deposits two
exposed to PBS fold compared to AD
alone [124]. mice injected with
PBS alone [124].
Pre-treatment of BV-
t
ip
2 microglia with In vivo studies of
MPL caused a two- ischemia using male
cr
fold increase in albino Wistar rats
internalized Aβ aged six to eight
us
compared to cells weeks.
exposed to Aβ alone
Results:
an
[124].
Intracerebroventricul
ar injection of MPL
48 h prior to an
M
ischemic insult (1)

decreased infarct
ed
size 3.5 fold; (2)

improved memory
and learning three to
pt
seven fold; (3)

increased IRF3,
ce
IFN-β, and
transforming growth
Ac
factor-β
concentrations in
whole brain
homogenates two
fold compared to
ischemic mice

injected with the
dimethyl sulfoxide
vehicle solution only
[142].
t
ip
Abbreviations: Aβ = amyloid-β peptide; AD = Alzheimer’s disease; APPSWE/PS1 = amyloid
cr
precursor protein/presenilin mice; BMDCs = bone marrow-derived dendritic cells; BV-2 =
immortalized murine microglia model; C57BL/6 = C57 black 6 mice; CD14 = cluster of
us
differentiation 14, IL = interleukin; KO = knock out, LPS = lipopolysaccharide; MD-2 =
myeloid differentiation factor 2; MIP = macrophage inflammatory protein; MPL =
monophosphoryl lipid A; N/A = not applicable since relevant data cannot be identified; PBMCs
an
= peripheral blood mononuclear cells; PBS = phosphate-buffered saline; TLR = toll-like
receptor; TNF = tumor necrosis factor
M
ed
pt
ce
Ac

Targeting Toll-Like Receptor 4 To Modulate Neuroinflammation in Central Nervous System Disorders

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Expert Opinion on Therapeutic Targets

ISSN: 1472-8222 (Print) 1744-7631 (Online) Journal homepage: https://www.tandfonline.com/loi/iett20

Targeting toll-like receptor 4 to modulate

To link to this article: https://doi.org/10.1080/14728222.2019.1676416

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Journal: Expert Opinion on Therapeutic Targets

Information Classification: General

Expert opinion: TLR 4 specific antagonists could suppress neuroinflammation by reducing

beneficial glial phagocytic activity with limited activity as inducers of pro-inflammatory

Key Words: Alzheimer’s disease, astrocytes, cytokines, microglia, neurodegeneration,

Information Classification: General

response protein 88 (MyD88)-independent pathway should be developed and further

Information Classification: General

Information Classification: General

Targeting TLR 4 with agonists or antagonists, or modulating its downstream signaling

pathogenesis of a range of neurological diseases, therapeutic agents targeting TLR 4 are

Information Classification: General

modulate their function if they are able to cross BBB.

We searched major databases including MEDLINE, PubMed Central, Science Citation

Information Classification: General

neurodegenerative diseases and other CNS pathologies.

Information Classification: General

Information Classification: General

Neuroinflammation is arguably the most destructive component of neurodegenerative

related DAMPs responsible for initiation of neuroinflammatory reactions in these pathologies.

In AD, the abnormal accumulation of Aβ contributes to neuroinflammation by directly

Information Classification: General

Information Classification: General

Information Classification: General

2.1.4. TLR 4 regulates oligodendrocytes and myelination

Demyelination of cerebral white matter is a neurodegenerative process that begins early

myelin by oligodendrocytes, thereby improving neuronal health. As MS progresses, myelin is

Information Classification: General

proteins. Autophagy is further classified into three categories referred to as micro-autophagy,

misfolding such as amyloidosis, synucleinopathies, and tauopathies.

2.2.1. TLR 4 mediates phagocytosis

Information Classification: General

Autophagy is a mechanism that facilitates removal of damaged organelles or abnormal

which could influence other TLR 4-mediated responses.

2.2.3. TLR 4 regulates apoptosis of CNS cells

Apoptosis is a hallmark of many CNS diseases. It is well established that apoptosis of

Information Classification: General

2.2.4. TLR 4 enhances Aβ protein clearance

TLR 4 activation enhances the phagocytosis of extracellular Aβ aggregates, which may

progression of cognitive symptoms [85,86]. Abnormal cleavage of amyloid precursor protein

Information Classification: General

α-Synuclein plays a key role in neurodegenerative conditions known as

The activation of TLR 4 may reduce aggregation of hyperphosphorylated tau (p-tau)

Information Classification: General

LPS, which is a component of the outer membrane of Gram-negative bacteria, over-activates

Information Classification: General

TAK-242, by blocking TLR 4 intracellular signaling, reduced symptoms of chronic

cytokines and NO.

Information Classification: General

3.2. Effects in the CNS

In addition, Gaikwad and Agrawal-Rajput [116] demonstrated that LPS-RS, a

neurotoxicity and inhibited microglial phagocytosis. Furthermore, immunofluorescence studies

Information Classification: General